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Article: Specific transbilayer translocation of dolichol-linked oligosaccharides by an endoplasmic reticulum flippase

TitleSpecific transbilayer translocation of dolichol-linked oligosaccharides by an endoplasmic reticulum flippase
Authors
Issue Date2009
PublisherNational Academy of Sciences. The Journal's web site is located at http://www.pnas.org
Citation
Proceedings Of The National Academy Of Sciences Of The United States Of America, 2009, v. 106 n. 3, p. 767-772 How to Cite?
AbstractThe oligosaccharide donor for protein N-glycosylation, Glc 3Man9GlcNAc2-PP-dolichol, is synthesized via a multistep pathway that starts on the cytoplasmic face of the endoplasmic reticulum (ER) and ends in the lumen where the glycosylation reaction occurs. This necessitates transbilayer translocation or flipping of the lipid intermediate Man5GlcNAc2-PP-dolichol (M5-DLO) across the ER membrane. The mechanism by which M5-DLO - or any other lipid - is flipped across the ER is unknown, except that specific transport proteins or flippases are required. We recently demonstrated M5-DLO flipping activity in proteoliposomes reconstituted from detergent-solubilized ER membrane proteins and showed that it was ATP-independent and required a trypsin-sensitive protein that sedimented at approximately 4S. By using an activity-enriched fraction devoid of glycerophospholipid flippase activity, we now report that M5-DLO is rapidly flipped in the reconstituted system with a time constant τ <2 min, whereas its triantennary structural isomer is flipped slowly with τ >200 min. DLOs larger than M5-DLO are also poorly translocated, with τ ranging from approximately 10 min to >200 min. We conclude that (i) the number and arrangement of mannoses in the DLO glycan has a profound effect on the ability of the DLO to be translocated by the flippase, (ii) glycan size per se does not dictate whether a DLO will be flipped, and (iii) the flippase is highly specific for M5-DLO. Our results suggest a simple structural model for the interaction between the DLO head group and the flippase. © 2009 by The National Academy of Sciences of the USA.
Persistent Identifierhttp://hdl.handle.net/10722/188679
ISSN
2015 Impact Factor: 9.423
2015 SCImago Journal Rankings: 6.883
ISI Accession Number ID
References

 

DC FieldValueLanguage
dc.contributor.authorSanyal, Sen_US
dc.contributor.authorMenon, AKen_US
dc.date.accessioned2013-09-03T04:12:44Z-
dc.date.available2013-09-03T04:12:44Z-
dc.date.issued2009en_US
dc.identifier.citationProceedings Of The National Academy Of Sciences Of The United States Of America, 2009, v. 106 n. 3, p. 767-772en_US
dc.identifier.issn0027-8424en_US
dc.identifier.urihttp://hdl.handle.net/10722/188679-
dc.description.abstractThe oligosaccharide donor for protein N-glycosylation, Glc 3Man9GlcNAc2-PP-dolichol, is synthesized via a multistep pathway that starts on the cytoplasmic face of the endoplasmic reticulum (ER) and ends in the lumen where the glycosylation reaction occurs. This necessitates transbilayer translocation or flipping of the lipid intermediate Man5GlcNAc2-PP-dolichol (M5-DLO) across the ER membrane. The mechanism by which M5-DLO - or any other lipid - is flipped across the ER is unknown, except that specific transport proteins or flippases are required. We recently demonstrated M5-DLO flipping activity in proteoliposomes reconstituted from detergent-solubilized ER membrane proteins and showed that it was ATP-independent and required a trypsin-sensitive protein that sedimented at approximately 4S. By using an activity-enriched fraction devoid of glycerophospholipid flippase activity, we now report that M5-DLO is rapidly flipped in the reconstituted system with a time constant τ <2 min, whereas its triantennary structural isomer is flipped slowly with τ >200 min. DLOs larger than M5-DLO are also poorly translocated, with τ ranging from approximately 10 min to >200 min. We conclude that (i) the number and arrangement of mannoses in the DLO glycan has a profound effect on the ability of the DLO to be translocated by the flippase, (ii) glycan size per se does not dictate whether a DLO will be flipped, and (iii) the flippase is highly specific for M5-DLO. Our results suggest a simple structural model for the interaction between the DLO head group and the flippase. © 2009 by The National Academy of Sciences of the USA.en_US
dc.languageengen_US
dc.publisherNational Academy of Sciences. The Journal's web site is located at http://www.pnas.orgen_US
dc.relation.ispartofProceedings of the National Academy of Sciences of the United States of Americaen_US
dc.subject.meshAnimalsen_US
dc.subject.meshBiological Transporten_US
dc.subject.meshEndoplasmic Reticulum - Metabolismen_US
dc.subject.meshGlycosylationen_US
dc.subject.meshLipid Bilayers - Metabolismen_US
dc.subject.meshPhospholipid Transfer Proteins - Physiologyen_US
dc.subject.meshPolyisoprenyl Phosphate Oligosaccharides - Metabolismen_US
dc.subject.meshProteolipids - Metabolismen_US
dc.subject.meshRatsen_US
dc.titleSpecific transbilayer translocation of dolichol-linked oligosaccharides by an endoplasmic reticulum flippaseen_US
dc.typeArticleen_US
dc.identifier.emailSanyal, S: sumana@wi.mit.eduen_US
dc.identifier.authoritySanyal, S=rp01794en_US
dc.description.naturelink_to_subscribed_fulltexten_US
dc.identifier.doi10.1073/pnas.0810225106en_US
dc.identifier.pmid19129492-
dc.identifier.scopuseid_2-s2.0-58849150433en_US
dc.relation.referenceshttp://www.scopus.com/mlt/select.url?eid=2-s2.0-58849150433&selection=ref&src=s&origin=recordpageen_US
dc.identifier.volume106en_US
dc.identifier.issue3en_US
dc.identifier.spage767en_US
dc.identifier.epage772en_US
dc.identifier.isiWOS:000262809700019-
dc.publisher.placeUnited Statesen_US
dc.identifier.f10001147275-
dc.identifier.scopusauthoridSanyal, S=16069600000en_US
dc.identifier.scopusauthoridMenon, AK=7202324192en_US

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